Name: ampliphox colorimetric detection on a dna microarray for influenza
Uploaded: 2011-06-09T12:00:00
Description: Watch this Scientific Journal Video about ampliPHOX Colorimetric Detection on a DNA Microarray for Influenza at JoVE.com

InDevR इस काम के वित्तपोषण के लिए NIH / U01AI070276 NIAID और R43AI077112 मानता है.

1. RT-पीसीआर का उपयोग नमूना प्रवर्धन <ol><li> नैदानिक ​​सामग्री या एक वायरल QIAcube स्वचालित न्यूक्लिक एसिड निकासी मंच के साथ संयोजन के रूप में Qiagen MinElute वायरस स्पिन किट का उपयोग कर से अलग है. वायरल शाही सेना से निकाल?...

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	<li>Kumar, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Widely+Used+Diagnostics+"DNA-Microarray"-A+Review.">The Widely Used Diagnostics "DNA-Microarray"-A Review.</a> Amer J Inf Dis. 5, 207-218 (2009).</li><li>Miller, M. B., Tang, Y. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Basic+Concepts+of+Microarrays+and+Potential+Applications+in+Clinical+Microbiology.">Basic Concepts of Microarrays and Potential Applications in Clinical Microbiology.</a> Clin Microbiol Rev. 22, 611-633 (2009).</li><li>Mikhailovich, V., Gryadunov, D., Kolchinsky, A., Makarov, A. 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ampliPHOX वर्णमिति का पता लगाने प्रौद्योगिकी यहाँ प्रस्तुत है एक तेजी से, कम घनत्व माइक्रोएरे अनुप्रयोगों के लिए एकल रंग प्रतिदीप्ति पता लगाने के लिए सस्ता विकल्प है. Schematically चित्र 1 में दिखाया गया है, पत...

<table border="1"><tbody><tr><th> / अभिकर्मक उपकरण </th><th> निर्माता </th><th> # सूचि </th><th> टिप्पणियाँ </th></tr><tr><td> Qiagen MinElute वायरस स्पिन किट </td><td> Qiagen </td><td> 57704 </td><td> एकल 60 μl elution </td></tr><tr><td> QIAcube </td><td> Qiagen </td><td> 9001292 </td><td> ऐच्छिक </td></tr><tr><td> अबी 9800 थर्मल Cycler फास्ट </td><td> एप्लाइड Biosystems </td><td> 4441166 </td><td></td></tr><tr><td> Qiagen OneStep RT-पीसीआर किट </td><td> Qiagen </td><td> 210210 </td><td> किट dNTPs उपयोग नहीं किया </td></tr><tr><td> 2x खोलना बफर </td><td> InDevR इंक </td><td> एमआई 5007 </td><td></td></tr><tr><td> Biotinylated dNTP मिक्स </td><td> InDevR इंक </td><td> एमआई ५,००९ </td><td></td></tr><tr><td> Lambda exonuclease </td><td> Epicentre Biotechnologies </td><td> LE032K </td><td> 2500 यू, 10U/μl </td></tr><tr><td> FluChip प्राइमर मिश्रण </td><td> InDevR </td><td> N / A </td><td> बिक्री के लिए अभी तक उपलब्ध नहीं है </td></tr><tr><td> कक्षीय शेखर </td><td> Madell प्रौद्योगिकी </td><td> ZD 9556-एक </td><td></td></tr><tr><td> डिब्बे धो </td><td> InDevR इंक </td><td> एमआई 4002 </td><td></td></tr><tr><td> रैक धो </td><td> InDevR इंक </td><td> एमआई 4003 </td><td></td></tr><tr><td> 2x संकरण बफर </td><td> InDevR इंक </td><td> एमआई 5004 </td><td></td></tr><tr><td> अंशांकन चिप्स </td><td> InDevR इंक </td><td> एपी ५००६ </td><td></td></tr><tr><td> Buffers ई. धो </td><td> InDevR इंक </td><td> एमआई 5005 </td><td></td></tr><tr><td> ampliRED </td><td> InDevR इंक </td><td> एपी 5004 </td><td></td></tr><tr><td> ampliTAG </td><td> InDevR इंक </td><td> एपी 5001 </td><td></td></tr><tr><td> 2x ampliTAG बफर </td><td> InDevR इंक </td><td> एपी 5002 </td><td></td></tr><tr><td> ampliPHY, ampliPHY बढ़ाने </td><td> InDevR इंक </td><td> एपी 5003 </td><td></td></tr></tbody></table>

ampliphox colorimetric detection on a dna microarray for influenza

DNA microarrays have emerged as a powerful tool for pathogen detection.1-5 For instance, many examples of the ability to type and subtype influenza virus have been demonstrated.6-11 The identification and subtyping of influenza on DNA microarrays has applications in both public health and the clinic for early detection, rapid intervention, and minimizing the impact of an influenza pandemic. Traditional fluorescence is currently the most commonly used microarray detection method. However, as microarray technology progresses towards clinical use,1 replacing expensive instrumentation with low cost detection technology exhibiting similar performance characteristics to fluorescence will make microarray assays more attractive and cost-effective.
The ampliPHOX colorimetric detection technology is intended for research applications, and has a limit of detection within one order of magnitude of traditional fluorescence11, with a main advantage being an approximate ten-fold lower instrument cost compared to the confocal microarray scanners required for fluorescence microarray detection. Another advantage is the compact size of the instrument which allows for portability and flexibility, unlike traditional fluorescence instruments. Because the polymerization technology is not as inherently linear as fluorescence detection, however, it is best suited for lower density microarray applications in which a yes/no answer for the presence of a certain sequence is desired, such as for pathogen detection arrays. Currently the maximum spot density compatible with ampliPHOX detection is &tilde;1800 spots/array. Because of the spot density limitations, higher density microarrays are not suitable for ampliPHOX detection.
Here, we present ampliPHOX colorimetric detection technology as a method of signal amplification on a low density microarray developed for the detection and characterization of influenza viruses (FluChip). Although this protocol uses the FluChip (a DNA microarray) as one specific application of ampliPHOX detection, any microarray incorporating biotinylated target can be labeled and detected in a similar manner. The microarray design and biotinylation of the target to be captured are the responsibility of the user. Once the biotinylated target has been captured on the array, ampliPHOX detection can be performed by first tagging the array with a streptavidin-label conjugate (ampliTAG). Upon light exposure using the ampliPHOX Reader instrument, polymerization of a monomer solution (ampliPHY) occurs only in regions containing ampliTAG-labeled targets. The polymer formed can be subsequently stained with a non-toxic solution to improve visual contrast, followed by imaging and analysis using a simple software package (ampliVIEW). The entire FluChip assay from un-extracted sample to result can be performed in about 6 hours, and the ampliPHOX detection steps described above can be completed in about 30 min.

Watch this Scientific Journal Video about ampliPHOX Colorimetric Detection on a DNA Microarray for Influenza at JoVE.com

ampliPHOX Colorimetric Detection on a DNA Microarray for Influenza

ampliPHOX वर्णमिति पहचान तकनीक के प्रोटीन के लिए प्रतिदीप्ति पता लगाने के लिए एक सस्ता विकल्प के रूप में प्रस्तुत किया है. Photopolymerization के आधार पर, ampliPHOX ठोस बहुलक बस कुछ ही मिनटों में नग्न आंखों को दिखाई धब्बे पैदा करता है. परिणाम तो imaged और एक सरल अभी तक शक्तिशाली सॉफ्टवेयर पैकेज के साथ स्वचालित रूप से व्याख्या की.

ampliPHOX इन्फ्लुएंजा के लिए एक डीएनए माइक्रोएरे पर वर्णमिति जांच

DNA microarrays have emerged as a powerful tool for pathogen detection.1-5  For instance, many examples of the ability to type and subtype influenza virus ...

Research

JoVE Journal

Immunology and Infection

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इन्फ्लुएंजा वायरस के लिए उच्च throughput जांच विधि

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